Type 1 Diabetes (T1D) is an autoimmune disease that targets insulin secreting pancreatic beta cells for destruction and remains a major cause of morbidity and mortality in over 1% of the population worldwide. Although insulin therapy and islet transplantation are currently the most effective treatments, both of these approaches suffer from major limitations including the inability of insulin therapy to totally prevent diabetes- associated complications, the limited availability of human islets for transplantation, and the rejection of islet allografts irrespective of the use of immunosuppressive drugs. Therefore, the use of islets from a different species, such as pigs, and induction of tolerance without the chronic use of immunosuppressive drugs will be important therapeutic advancements required for the efficient treatment of T1D in the clinic. The main objective of this proposal is to establish proof-of-principle for a novel, safe, and practical approach designated as ApoFasL to induce transplantation tolerance to xenogeneic pancreatic islets as a therapeutic intervention for the treatment of Type 1 diabetes Given the important role played by Fas/FasL system in immune tolerance to self antigens, immune privilege, and lymphocyte homeostasis, and the extensive use of FasL to induce allograft tolerance in various settings, the Principle Investigator of this grant, Dr. Haval Shirwan, developed a novel form of FasL with potent apoptotic activity using ApoImmune's ProtEx(tm) technology. ProtEx(tm) involves the generation of chimeric immunological ligands with a modified form of core streptavidin, modification of a cell membrane with biotin, and decoration of biotinylated membrane with chimeric proteins. Preliminary data have demonstrated that the display of FasL on pancreatic islets (ApoFasL therapy) using ProtEx(tm) resulted in the prevention of graft rejection and treatment of chemically-induced diabetes in an allogeneic transplant model. Expanding on these data, the focus of this proposal is to develop proof-of-principle for an effective, clinically applicable approach to induce immune tolerance to xenogeneic islet grafts in a rat-to-mouse model for the treatment of diabetes. FasL will be displayed on rat donor islets for transplantation into both chemically-induced diabetic mice and spontaneously diabetic non-obese diabetic (NOD) mice, which exhibit destructive autoimmune pancreatic insulitis. Transplants will be done in the presence of rapamycin a pharmaceutical agent used in the clinic for the prevention of graft rejection. We hypothesize that islets decorated with FasL will induce tolerance by eliminating xenoreactive lymphocytes via apoptosis and expanding T regulatory cells. Rapamycin is expected to further augment this response by eliminating T effector cells and/or expanding T regulatory cells, thereby creating donor specific immune tolerance, leading to the survival of transplanted islets. Proof-of-principle will be followed by a Phase II STTR application to further develop ApoFasL into a lead product by testing its efficacy in a pig-to-nonhuman primate xenograft model. Tolerance to xenogeneic islets will serve as the first step towards the translation of this novel approach to the clinic for the treatment of T1D. If proven effective, ApoFasL may improve the quality of life of millions of individuals worldwide with an economic impact in billions of dollars. Xenograft islet transplantation represents an alternative therapeutic approach for Type 1 diabetes over classic allogeneic islet transplantation and insulin treatment therapies, and promises to yield a more definitive solution. The ApoFasL approach, when successful, will ensure a pathway to the treatment of Type 1 diabetes. [unreadable] [unreadable] [unreadable]